JPH1149675A - Prophylactic and treating agent for allergic disease containing compound ab5366 and enzyme inhibitor - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a prophylactic and treating agent useful as an antiallergic diseases and an enzyme inhibitor, which is effective to prevent/treat allergic diseases, and has low toxicity and new chemical structure different from conventional antiallergic agents. SOLUTION: The medicine is obtained by including compound AB5366 shown by the formula or a salt thereof as an active ingredient. They serve as an enzyme inhibitor against a synthetic enzyme catalyzing conversion of leukotriene A4 leukotriene C4 , leukotriene A4 hydrolase, or 5-lipoxygenase.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a pharmaceutical composition which is useful as a therapeutic or preventive drug for various allergic diseases caused by allergy in humans and animals, and which contains an AB5366 substance or a salt thereof as an active ingredient. Also,
The present invention relates to an enzyme inhibitor comprising an AB5366 substance or a salt thereof.

[0002]

2. Description of the Related Art In type I allergic reactions classified by Gell and Coombs, exogenous antigens bind to IgE on the receptor.
In response to (in the case of humans), so-called anaphylactic eel responsiveness of leukotrienes C ₄ , D ₄ , E ₄ among chemical mediators released from cells such as activated mast cells and leukocytes in response thereto Substance (slow reacti
It is already well known that ngsubstance of anaphylaxis (abbreviation: SRS-A) has a profound causal relationship with respect to the development of allergic diseases such as bronchial asthma and anaflexia. Dahlen, SE et al. Tested the muscle contractile activity of leukotriene, histamine and prostaglandin on bronchial muscles isolated from pollen sensitive patients.
Among them, leukotriene was found to show about 1000 times stronger muscle contraction activity than other substances. In the Schult-Dale test using this antigen and the isolated bronchial muscle, the action of an antihistamine on the bronchial muscle was ineffective,
It has been reported that the U-60257 substance, a leukotriene C ₄ antagonist, almost completely inhibited muscle contraction by an antigen [“Metabolism”, Vol. 20, No. 10, pp. 1195-1206 (1983)].

Leukotrienes B _4, the degranulation for leukocytes in low concentrations, vascular permeability enhancing, migration increased, are known to have a biological activity of causing calcium transport and the like. The inflammation is thought to be due to the action of leukotriene B _4, for example, rheumatism, spinal arthritis, gout, psoriasis, ulcerative colitis, there is a respiratory disease (JP-A-8-143529
issue). Leukotriene B ₄ antagonists are used for chronic rheumatoid arthritis, inflammatory bowel disease, psoriasis, nonsteroidal anti-inflammatory drug-induced gastric disease, adult respiratory distress syndrome, myocardial infarction, allergic rhinitis, hemodialysis-induced neutropenia, late stage It is known that it is useful for treating asthma (Japanese Patent Application Laid-Open No. 5-239008).

The biosynthetic pathway from arachidonic acid to leukotrienes or hydroxyeicosatetraenoic acids is as follows:
It is performed through the enzymatic action of 5-lipoxygenase.
Therefore, it is well known that 5-lipoxygenase inhibitors are effective in treating various inflammations by suppressing the production of chemical mediators [“Proteins, Nucleic Acids, Enzymes”, Vol. 33, No. 2, 136. 149 (1988)].

To date, biochemical and pharmacological studies on leukotrienes have been advanced, but their metabolic pathways and mechanisms of allergic development are complicated, and many points remain unclear. Therefore, the effects of the antiallergic drugs developed so far have not always been sufficient in clinical practice. In addition, there are many substances that are effective as anti-allergic drugs but have not been put to practical use because of their high toxicity.

There are very few substances produced by microorganisms that inhibit the biosynthesis system of leukotriene, and recently, the substances produced by actinomycetes (Streptomyces diastaticus) 8
(S) - amino -2 (R) - ".. J. Nat Prod" methyl-7-oxo-nonanoyl dichroic acids are known to inhibit the biosynthesis of the leukotrienes A ₄ to B ₄ [Vol. 59 No. 10, 962
-964 (1996).

[0007] Leukotriene A ₄ to Leukotriene C
Substances that inhibit or suppress the biosynthesis of ₄ include acetylenic acids such as 4,7,10-hexadecatriic acid, prostacyclin derivatives, U-60257 substances, and intracellular glutathione depleting agents. And diethyl maleate, buthionine sulfoxide and the like are known.

[0008] On the other hand, AB5366 substance is an antibiotic known as a medical antifungal agent and an agricultural and horticultural fungicide (Japanese Patent Application Laid-Open No. 8-242873). This AB5366 substance is produced by cultivation of AB5366 strain belonging to the genus Pikunidiera (deposited with the National Institute of Bioscience and Human Technology, FERM P-14662), and is a pale yellow powder soluble in water, methanol, ethanol, and dimethyl sulfoxide. And its molecular formula is C ₁₉ H ₃₇ NO ₈ S (molecular weight 439), and its chemical structure is a substance represented by the following formula (I).

[0009]

SUMMARY OF THE INVENTION
Currently, it is demanded to obtain a novel anti-allergic agent which can suppress the type of allergic reaction and has low toxicity.
In particular leukotriene A ₄ hydrolase involved in the biosynthetic conversion of enzyme and leukotriene A ₄ involved in the biosynthetic conversion of leukotriene A ₄ to leukotriene C ₄ to leukotriene B _4, and the arachidonic acid 5-hydroxy Eicosatetraenoic acid (5-HPE
There is a need for novel enzyme inhibitors that can inhibit the enzymatic activity of 5-lipoxygenase involved in the biosynthetic pathway to TE).

An object of the present invention is to provide a novel and useful drug having desirable properties as an excellent therapeutic or preventive agent for allergic diseases, which can satisfy the above-mentioned demand. It is intended to solve the above-mentioned problems associated with the prior art.

[0011]

Means for Solving the Problems In order to achieve the above object, the present inventors have studied a bioactive substance produced by a microorganism, which has an inhibitory effect on the enzymes of the biosynthesis of leukotrienes B ₄ and C _4. Intensive research has been conducted to find a physiologically active substance which has an inhibitory action on the enzyme activity of and / or 5-lipoxygenase and has an entirely new chemical structure as an antiallergic agent. As a result, it said AB5366 substances known to have antifungal activity, biosynthesis from leukotriene A ₄ to leukotriene C _4,
Also biosynthesis from leukotriene A ₄ to leukotriene B _4, was found to further inhibit various enzymes involved in biosynthesis of arachidonic acid to 5-hydroxy eicosatetraenoic acid (5-HPETE).

The AB5366 substance comprises leukotriene and 5-HP
Inhibiting ETE biosynthesis is the first discovered fact. The AB5366 substance is a substance having a novel structure which is clearly different from conventionally known anti-allergic agents. When the AB5366 substance is administered, it has the above-mentioned enzyme inhibitory activity, so that leukotrienes C ₄ , D ₄ ,
Since E ₄ and B ₄ are not produced, it can be expected to be effective for prevention or treatment of allergic diseases. Based on these findings, the present invention has been completed.

Therefore, according to the first aspect of the present invention, the following formula (I) Or a pharmaceutically acceptable salt thereof represented by the formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient.

Examples of salts of the AB5366 substance include salts with pharmaceutically acceptable organic bases, such as quaternary ammonium salts, and salts with various pharmaceutically acceptable metals, such as alkali metal salts and There are alkaline earth metal salts and ammonium salts. It can be in the form of a mono or disalt. Also, pharmaceutically acceptable acids include so-called mineral acids such as hydrochloric acid and sulfuric acid, and acid addition salts with organic acids such as acetic acid and caproic acid.

The AB5366 substance has low toxicity, and when it is used as a medical active ingredient, it can be administered orally or parenterally.

The first drug of the present invention can be used as an injection, an oral preparation, an ointment, a suppository, etc. in the form of a composition containing the AB5366 substance or a salt thereof as an active ingredient alone or in combination with an excipient. Can be formulated. The excipient can be a pharmaceutically acceptable liquid carrier, such as water or ethanol, or a solid carrier, such as starch. The type and composition of the carrier are appropriately selected depending on the administration route and administration method. The dose of the AB5366 substance or its salt varies depending on age, body weight, etc., but is usually 1 mg to 1,000 m for adults.
About g can be administered orally or parenterally (for example, by intravenous injection).

In the second invention, the above formula (I)
Characterized in that it consists AB5366 substance or a pharmacologically acceptable salt thereof in, provides enzyme inhibitors against biosynthesis enzyme or leukotriene A ₄ hydrolase or 5-lipoxygenase from leukotriene A ₄ to leukotriene C ₄ Is done.

The enzyme inhibitor according to the second aspect of the present invention is useful for academic research on various enzymes in animal organisms, biochemical research for allergic diseases, and biochemical analysis or diagnosis of blood samples.

Next, AB53 used as an active ingredient in the present invention.
The fact that 66 substances have enzyme inhibitory activity against various enzymes is illustrated in Test Examples 1 to 3 described below.

The enzyme inhibitory activity test leukotriene C ₄ for biosynthetic enzymes Test Example 1 leukotriene C ₄ is biosynthesized by glutathione to the carbon of a 6-position of the leukotriene A ₄ in the body is added by the action of the enzyme However, the AB5366 substance used in the present invention has an effect of inhibiting this biosynthetic process. For the assay of the enzyme inhibitory activity of AB5366 against leukotriene C ₄ biosynthetic enzyme, a paper by Bach MK et al.
Pharmacol., 34, 2695-2704 (1985)] and Fitz
Patric F.A. et al. [“J. Biol. Chem.” 257 4680-
4683 (1982)].

The biosynthetic enzymes of leukotriene C ₄ used for the test was used crude enzyme extract fraction obtained from basophilic leukemia cells of the rat (RBL-1).

In the reaction system, leukotriene A ₄ methyl ester as a substrate, albumin as a stabilizer,
The final concentration of the AB5366 substance was 1000, 300 with respect to the mixture of the serine / borate complex for preventing conversion of the formed leukotriene C ₄ to D ₄ , the crude enzyme extract fraction, and the buffer. , 100, 30 or 10 μM of each concentration of the AB5366 substance solution was added. These reaction mixtures were reacted at 37 ° C. for 15 minutes to advance the enzyme reaction. Thereafter, ice-cooled methanol was added to stop the enzyme reaction. The resulting product was quantified of leukotriene C ₄ in the reaction solution was carried out by radioimmunoassay (RIA).

[0023] After performing the enzyme reaction as above without addition of AB5366 substance as a control test to measure the produced amount of leukotriene C ₄ in the reaction solution of the control test. Relative to the amount of leukotriene C ₄ generated in this control test to measure the produced amount of leukotriene C ₄ in each test described above using AB5366 material, inhibition of leukotriene C ₄ production by AB5366 substance at each concentration ( %) Was calculated. The results obtained are shown in Table 1 below.

[0024]

From the above results shown in Table 1, the IC ₅₀ value of the enzyme inhibitory activity of AB5366 substance against the biosynthetic enzyme of leukotriene C ₄ was calculated. The results are shown in Table 2 below.
Incidentally, as described in Table 2 diethylcarbamazine and IC ₅₀ of sulfasalazine as a positive control reference value.

[0026]

Test Example 2 Test for enzyme inhibitory activity of leukotriene B ₄ on biosynthetic enzyme In the biosynthetic conversion of leukotriene C ₄ to leukotriene B ₄ in a living body, leukotriene A ₄ hydrolase is involved as a biosynthetic enzyme. Test of enzyme inhibition activity of AB5366 agent on leukotriene A ₄ hydrolase, Radmark, O. et al. [LEUKOTRIENE A ₄ hydrolase
in human leukocytes, "J. Biol. Chem." 259 12339
−12345 (1984)) and a paper by Kuhl, P. et al. (Ebselen reduc
es theformation of LTB ₄ in human leukocytes by iso
merisation to its 5S, 12R-6-trans-isomer, `` Prosta
glandins, Vol. 31, pp. 1029-1048 (1986)) and Izumi, T.
Tissue distribution of leukotriene A ₄ hy
drolase activity inguinea pig, "Biochem. Biophys.
Res. Comm., 135, 139-145 (1986)].

That is, the reaction system used was a reaction mixture consisting of leukotriene A ₄ hydrochloride, leukotriene A ₄ hydrolase, a buffer, and AB5366 substance added at a predetermined concentration as a substrate. After reacting at 37 ° C. for 1 minute, ice-cooled methanol was added to the reaction solution to stop the enzyme reaction. It was quantified by further generated leukotriene B ₄ radioimmunoassay (RIA). From the measured value of the amount of leukotriene B ₄ produced, the concentration of the AB5366 substance that inhibits the activity of leukotriene A ₄ hydrolase by 50% (IC ₅₀ value) was calculated. In addition, the same test was performed using ebselen as a positive control. The results are shown in Table 3 below.

[0029]

Test Example 3 Test for enzyme inhibitory activity on 5-lipoxygenase This test was carried out by Shimuzu, T. et al. [Enzyme with dual lipo
xygenase activitiescatalyzes leukotriene A ₄ synthe
tase from arachidonic acid, "Proc. Natl. Acad. Sc
i., USA, 81, 689-693 (1984) '' and Egan, RW,
And a paper by Gale, PH [Inhibition of mammalian 5-
lipoxygenase by aromatic disulfides. "J. Biol. Che
m. "260, 11554-11559 (1985)].

That is, 5-lipoxygenase used as a test enzyme was prepared from rat basophil leukemia cells (RBL-1). Various concentrations of the AB5366 substance and 5-lipoxygenase were added to the buffer and pre-incubated for 5 minutes at room temperature. Thereafter, arachidonic acid as a substrate was added to the mixture and reacted for 8 minutes. Thereafter, citric acid was added to stop the enzyme reaction. Generated 5-hydroxyeicosatetraenoic acid (5-H
PETE) was quantified by radioimmunoassay (RIA).

[0032] From the measured values of the amount of 5-HPETE, the enzymatic activity of 5-lipoxygenase was calculated concentrations of AB5366 substance which inhibits 50% (IC ₅₀ value). As a positive control, BW-7
The same test was performed using 55C substance and phenidone. The results are shown in Table 4 below.

[0033]

Test Example 4 Acute Toxicity Test (a) Using an ICR male mouse (5 weeks old), an acute toxicity test was conducted by oral administration of AB5366 substance (mono-Na salt). LD ₅₀ values of AB5366 substance upon oral administration was low toxicity shows the above 500 mg / kg. Since its LD ₅₀ value was higher than the effective amount required for inhibiting leukotriene C ₄ biosynthetic enzyme, its safety was confirmed.

(B) Using an ICR male mouse (6 weeks old), an acute toxicity test was performed by intravenous administration of AB5366 substance (mono-Na salt). LD ₅₀ value of AB5366 substance at the time of intravenous injection is, 10
The toxicity was low, indicating 0 mg / kg or more. High safety was confirmed by intravenous injection of AB5366 substance or its salt.

[0036]

According to the present invention, there is provided a prophylactic / therapeutic agent for an allergic disease which comprises a highly safe AB5366 substance or a salt thereof as an active ingredient, having a novel chemical structure as an antiallergic agent. .

Claims (2)

[Claims] 1. The following formula (I) A preventive or therapeutic agent for an allergic disease, comprising an AB5366 substance represented by the formula (I) or a pharmaceutically acceptable salt thereof as an active ingredient: 2. The following formula (I) Enzyme inhibitors on the AB5366 substance or characterized in that it consists a pharmaceutically acceptable salt thereof, biosynthetic enzymes or leukotriene A ₄ hydrolase or 5-lipoxygenase from leukotriene A ₄ to leukotriene C ₄ are represented in.